Studies have now revealed that Alzheimer's disease (AD) patients have more severe atherosclerosis than age-matched controls without AD, and established a positive correlation between the degree of atherosclerotic disease of cerebral arteries in AD patients and neuropathological features typical for AD. However, mechanisms by which cholesterol (CL) metabolism influences AD pathogenesis remain uncertain. CL efflux and generation of HDL are mediated by ABCA1 membrane transporter which is transcriptionally controlled by Liver X nuclear Receptors (LXR). ABCA1 mutations cause severe HDL deficiencies, like hypoalphalipoproteinemia and Tangier Disease (TD). A major goal of our research is to reveal the role of ABCA1 in the molecular pathogenesis of AD and thus to explore and to test new therapeutic strategies. The central hypothesis is that ABCA1 affects ABeta formation/deposition and clearance. In support are our studies (currently in press) funded by NIA R21 and R03 awards, showing that ABCA1 deficiency in APP23 mice leads to a dramatic decrease of soluble apoE and an increased deposition of ABeta in the brain. We have also demonstrated that application of the synthetic LXR ligand T0901317 (TO) in vivo increases the expression of ABCA1 in CNS and decreases the amounts of ABeta species in the brain. We found that primary cells from TD patients generate and secrete more ABeta and that their response to LXR synthetic ligands is different and depend on the type of ABCA1 mutation. Importantly, one of the primary ABCA1 mutant cell lines with a point mutation causing N935S amino acid substitution was established from a TD patient with dementia and abundant amyloid deposits in the brain, but without cardiovascular pathology. Thus, specific mutations in ABCA1 differentially disturb its regulatory role on beta-amyloid deposition and atherosclerosis, and therefore we hypothesize that ABCA1 mutations determine the clinical phenotype observed in patients by at least partly different mechanisms. The goal of this NIA pilot grant is to develop and characterize a mouse line that expresses mutated ABCA1N935S. Aim 1. To create genetically engineered mice that express mutated ABCA1N935S. This mouse line will be created by utilizing a "Knock-in" approach to introduce an N935S mutation in the ABCA1 locus in mouse ES cells. These ABCA1S/S mice will ubiquitously express ABCA1N935S, the mutated gene will remain under the endogenous ABCA1 transcriptional control and the mutant protein will replace wild type ABCA1. Aim 2. To characterize ABCA1N935S knock-in mice. We will analyze in vivo the expression of ABCA1N935s and its function as a cholesterol transporter by measuring plasma CL and lipoproteins. The effect of the mutated ABCA1N93ss on brain lipoproteins and endogenous APR processing and ABeta generation in vivo will be determined as a preliminary step to substantiate future detailed biochemical studies. [unreadable] [unreadable] [unreadable]